Soybean variety BK000496

ABSTRACT

Disclosed is the seed of a novel soybean variety, designated BK000496, a sample of which is deposited under ATCC Accession No. PTA-122489. Also disclosed are plants, or parts thereof, grown from the seed of the variety, plants having the morphological and physiological characteristics of the BK000496 variety, and methods of using the plant or parts thereof in a soybean breeding program.

CROSS-REFERENCE TO RELATED APPLICATION

This application claims priority to U.S. Provisional Patent ApplicationSer. No. 61/908,331, filed Nov. 25, 2013, the disclosure of which ishereby incorporated herein in its entirety by this reference.

FIELD OF THE INVENTION

This invention relates generally to the field of soybean breeding. Inparticular, the invention relates to a soybean variety designatedBK000496 that includes plants, plant parts, and seeds of soybean varietyBK000496. Methods for producing soybean plants by crossing soybeanvariety BK000496 with itself or any different soybean plant are anintegral part of this invention as are the resultant soybean plants,including the plant parts and seeds. This invention further relates tomethods for producing BK000496-derived soybean plants and to methods forregenerating such plants from tissue cultures of regenerable cells aswell as the plants obtained therefrom. Methods for producing a soybeanplant containing in its genetic material one or more transgenes and tothe transgenic soybean plants produced by that method are also a part ofthis invention.

BACKGROUND OF THE INVENTION

Soybean (Glycine max) is a major grain crop valued for the high levelsof oil and protein found in soybean seed. Soybean breeding has resultedin significant improvements in yield potential, stability of yield,adaptation of the species to mechanical harvest, and yield protectionthrough improved disease resistance.

Due to the nature of plant science agriculture, broadly defined as amanipulation of available plant resources to meet the needs of thegrowing human population, the environment in which plants are grown foragricultural production continuously offers new obstacles toagricultural production. Each new variety, or cultivar, released toagricultural production is selected for the purpose of increasing yieldresulting from increased disease resistance to prevalent diseases, orfrom direct or indirect improvement in yield potential or efficiency ofproduction. Development of stable, high yielding varieties with superiorcharacteristics is an ongoing goal of soybean breeders.

There is a need in the art for a novel, superior soybean variety andsoybean seed.

SUMMARY OF THE INVENTION

In one aspect, the present invention provides a soybean seed designatedBK000496, wherein a sample of said seed is deposited under ATCCAccession No. PTA-122489.

In another aspect, the present invention provides a soybean plant, or apart thereof, produced by growing seed designated BK000496, or a soybeanplant having all the physiological and morphological characteristics ofthe soybean plant when grown in the same environmental conditions, or apollen grain, an ovule, a protoplast, a cell, an embryo, a cotyledon, ahypocotyl, a meristem, a root, a pistil, an anther, a flower, a stem, apod, a leaf, or a petiole of a soybean plant according to the presentinvention.

In yet another aspect, the present invention provides a tissue cultureof protoplasts or regenerable cells from a plant, or parts thereof,produced by growing seed designated BK000496, and a soybean plantregenerated from the tissue culture.

In still another aspect, the present invention provides a method forproducing a soybean seed, and soybean seed produced by the method, aswell as plants grown from seed produced by the method are provided. Themethod comprises crossing soybean plants and harvesting the resultantseed, wherein at least one soybean plant is the soybean variety BK000496of the present invention.

In another aspect, a method for producing a soybean varietyBK000496-derived soybean plant, and soybean variety BK000496-derivedsoybean plants, or parts thereof, produced by the methods are provided.The method comprises crossing a soybean variety BK000496 plant of thepresent invention with a second soybean plant to yield progeny soybeanseed and growing the progeny seed to yield a soybean varietyBK000496-derived soybean plant. In some embodiments, the method furthercomprises crossing the soybean variety BK000496-derived soybean plantwith itself or another soybean plant to yield an additional soybeanvariety BK000496-derived soybean progeny seed and growing this progenysoybean seed to yield additional soybean variety BK000496-derivedsoybean plants. In some embodiments, these last two steps are repeatedat least one time to generate additional soybean varietyBK000496-derived soybean plants.

In yet another aspect, a method for producing a plant of soybean varietyBK000496 comprising an added desired trait, and plants produced by themethod, are provided. In some embodiments, the method comprisesintroducing at least one transgene or locus conferring the desired traitinto the soybean variety BK000496 plant. In certain embodiments, thedesired trait is selected from the group consisting of male sterility,site-specific recombination, abiotic stress tolerance, herbicidetolerance, insect or pest resistance, disease resistance, fungalresistance, modified fatty acid metabolism, modified protein metabolism,and modified carbohydrate metabolism. In other embodiments, the desiredtrait is herbicide tolerance and the tolerance is conferred to one ormore herbicides selected from the group consisting of glyphosate,phenoxyacetate auxins (such as 2,4-diclorophenoxyacetic acid (2,4-D)),pyridyloxyacetate auxins (such as fluroxypyr and triclopyr),phenoxyproprionate auxins (such as MCPA and dichloprop),phenoxybutanoate auxins (such as 2,4-DB), sulfonylurea, imidazalinone,dicamba, glufosinate, cyclohexone, triazine, and benzonitrile. In stillother embodiments, the desired trait is insect resistance and thetransgene encodes a Bacillus thuringiensis (Bt) endotoxin.

In still another aspect, a method of producing a progeny soybean varietyderived from variety BK000496 comprising a desired trait, and plantsproduced by the method, are provided. In some embodiments, the methodcomprises crossing a soybean variety BK000496 plant of the presentinvention with a plant of another soybean variety that comprises adesired trait to produce F1 progeny plants, selecting one or more F1progeny plants that have the desired trait to produce selected progenyplants, crossing the selected progeny plants with the BK000496 plants toproduce backcross progeny plants, selecting for backcross progeny plantsthat have the desired trait and physiological and morphologicalcharacteristics of soybean variety BK000496 to produce selectedbackcross progeny plants, and repeating the last two steps a sufficientnumber of times in succession to produce selected second or higherbackcross progeny plants that comprise the desired trait and thephysiological and morphological characteristics of soybean varietyBK000496 when grown in the same environmental conditions. In someembodiments, the last two steps are repeated three or more times insuccession to produce selected fourth or higher backcross progenyplants. In some embodiments, the desired trait is selected from thegroup consisting of male sterility, herbicide resistance, insectresistance, modified fatty acid metabolism, modified carbohydratemetabolism and resistance to bacterial disease, fungal disease or viraldisease.

In yet another aspect, a method of producing a commodity plant productis provided, which comprises obtaining a plant of the present invention,or a part thereof, and producing the commodity plant product therefrom.In some embodiments, the commodity plant product is soybean meal, soyflour, soy protein concentrate, soy protein isolates, texturized soyprotein concentrate, hydrolyzed soy protein, whipped topping, whole soyseed, processed soy seed, animal feed, soymilk, soy nut butter, natto,tempeh, edible raw whole soybean pods, roasted whole soybean pods,edamame, soymilk, soy yogurt, soy cheese, tofu, yuba, and biomass.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

In the claims, descriptions, and tables that follow, numerous terms areused and are defined as follows:

Canopy width is measured at the widest point of the soybean plant.Visually scored from 1 to 9, where 1=very narrow canopy and 9=extremelybushy canopy.

Cotyledon is a type of seed leaf. The cotyledon contains the foodstorage tissues of the seed. Cotyledon color can be measured as acharacteristic of a variety.

Firmness is defined as the force necessary to attain a given deformationof the tofu, applied either by compression or plunger penetration andexpressed in Newtons per mm.

Flower color: Modern soybeans are characterized by two major flowercolors, purple or white. Some varieties are heterogeneous for flowercolor whereby some plants have purple flowers and some have white.

Growth habit refers to stem termination in soybeans and the resultantdifferences in flower production. “Indeterminate” varieties continue togrow during the reproductive phase, producing new branches and nodesafter flowering is well underway. “Determinate” varieties tend to delaythe onset of flowering somewhat, and limit new node and branchdevelopment after flowering has been initiated. “Semi-determinate”varieties continue to produce new vegetative growth during thereproductive phase but growth terminates more quickly than inindeterminate varieties.

Hardness is defined as the force necessary to attain a given deformationof the tofu, applied either by compression or plunger penetration andexpressed in Newtons.

Hilum refers to the point of attachment of soybean seed to maternaltissue.

Hilum color in modern soybeans may be black, brown, yellow, gray, buff,or imperfect black.

Leaflet shape: The leaflet may be broad or narrow and may be lanceolate,ovate or oval in shape.

Lodging relates to the stature of the plant relative to the ground.Lodging resistance is rated on a scale of 1 to 5. A score of 1 is givento an erect plant (lodging resistant). A score of 3 is given to a plantthat is leaning at a 45-degree angle relative to the ground. A score of5 indicates a plant lying on the ground.

Maturity date is the date when 95% of pods have turned color from greencolor to their mature brown or tan color. The maturity date is countedin days and is calculated from January 1.

Maturity group refers to an industry division of groups of varietiesbased on the zones in which the varieties are adapted. Soybeans maturedifferentially in response to day-length and thus to latitude wheregrown. In the soybean production areas of the United States, forexample, the northernmost production region of northern Minnesota isplanted to soybeans that mature under very long day-lengths during earlysummer. In the southernmost production regions of the Southeast,soybeans that mature from the influence of short day-length during earlysummer are grown. Those adapted to northern day-lengths are classifiedas early-maturing, those adapted to the southern regions are classifiedas late-maturing. Maturity groups include very long day length varieties(000, 00, 0) and extend to very short day length varieties (VII, VIII,IX, X). For example, maturity group I soybean varieties are typicallygrown in southern Minnesota, whereas maturity group IV soybean varietiesare typically group in southern Illinois.

Plant includes plant cells, plant protoplasts, plant cell tissuecultures from which soybean plants can be regenerated, plant calli,plant clumps, and plant cells that are intact in plants or partsthereof. “Plant part” includes, but is not limited to, embryos,protoplasts, cells, pollen, ovules, cotyledons, hypocotyls, meristems,roots, pistils, anthers, flowers, stems, leaves, pods, petioles, and thelike.

Plant height is measured from the top of soil to top node of the plantin any convenient unit of length (i.e., inches, centimeters, etc.). Forthe data presented herein, plant height was measured just prior toharvest and is expressed in centimeters.

Pod wall color refers to the color of the mature pod wall, as distinctfrom the color of the pubescence, and in modern soybeans, may be brownor tan.

Pubescence relates to the plant trichomes or hairs found on the stems,leaves and pods of soybeans.

Pubescence color in modern soybeans may be tawny, gray or light tawny.

Relative maturity: Within maturity groups, a more precise maturityassignment is given that subdivides each maturity group into tenths. Forexample, a relative maturity of 3.3 is assigned to a late early maturitygroup III soybean variety.

Roundup Ready 2 Tolerance the transgenic soybean event MON89788 fromMonsanto Company, as identified in the USDA petition extension No06-1780-01p, which imparts glyphosate tolerance to the plant.

Seed coat color refers to the color of the seed coat, and in modernsoybeans may be yellow, green, brown or black.

Seed coat luster refers to the luster of the seed coat, and may be dullor shiny.

Seed coat peroxidase activity is defined by a chemical taxonomictechnique to separate varieties based on the presence or absence of theperoxidase enzyme in the seed coat. There are two types of soybeanvarieties, those having high peroxidase activity and those having lowperoxidase activity. Ratings are HIGH or LOW for peroxidase enzymeactivity.

Seed size is measured by seed number per pound of seed. Seed size is aheritable trait but is influenced by environment, and as such, is oftenpresented as a comparison to another variety.

Shattering refers to pod dehiscence prior to harvest resulting in a lossof mechanically harvestable seed. Pod dehiscence involves seeds fallingfrom the pods to the soil. This is visually scored with a 1 to 9 scalecomparing all genotypes within a given test. A score of 1 means podshave not opened and no seeds have fallen out. A score of 5 indicatesapproximately 50% of the pods have opened, with seeds falling to theground and a score of 9 indicates 100% of the pods are opened.

Solids content of soymilk refers to the amount of solids in soymilkdivided by solids soybean and multiplied by 100%.

Soybean emergence scores, also referred to simply as “Emergence,” ratethe ability of the seedlings to emerge from the soil. A visual score of1 to 9, taken from emergence to V3, is used whereby a score of 1 to 3indicates excellent emergence vigor and early growth, an intermediatescore of 5 indicates average ratings, and a score of 7 to 9 indicates avery poor emergence vigor and early growth.

Soymilk yield refers to the yield of soymilk expressed as liters per kgof protein or liters per kg of bean.

Sulfonylurea Tolerance is a herbicide-tolerance trait that improvessoybean tolerance to ALS (acetolactate synthase) inhibitor herbicides.Sulfonylurea Tolerant soybeans carry a modified ALS gene, which enhancesthe variety's natural tolerance to sulfonylurea.

Water Uptake is measured by soaking soybeans in water and recording theweight increase in beans with respect to time.

Yield refers to the yield of seed harvested from a soybean crop. Yielddata presented herein is expressed as bushels of seed/acre and is theactual yield of the grain at harvest.

Soybean Variety BK000496

The present invention provides plants, seeds, plant parts, andderivatives thereof of the soybean variety BK000496, characterized bymolecular and physiological data obtained from the representative sampleof said variety deposited with the American Type Culture Collection(ATCC). The present invention further provides methods for producingsoybean variety BK000496 and methods for breeding with soybean varietyBK000496 to produce novel derived soybean varieties.

Soybean variety BK000496 has superior characteristics and was developedfrom crossing two elite soybean varieties. Some of the criteria used toselect the variety in various generations included seed yield, lodgingresistance, emergence, disease resistance and tolerance, herbicidetolerance, maturity, late season plant intactness, plant height, andshattering resistance. The breeding history of the variety is summarizedin Table 1.

TABLE 1 Breeding Method for Cultivar BK000496 Filial Generation MethodF0 cross between parents F1 plant growout F2 population growout F3progeny row, single-plant selection F4 plant-row yield trial F5preliminary yield trial F6 purity reselection and seed increase F7advanced yield trial F8 seed increase F9 advanced yield trial

Soybean variety BK000496 has excellent agronomic characteristicsincluding high yield potential relative to lines of similar maturity.Soybean variety BK000496 is well-adapted to early maturity group 0 tomid group 0 growing areas of Michigan, Minnesota, North Dakota, SouthDakota, and Wisconsin.

Soybean variety BK000496 has been judged to have uniformity andstability of its morphological and other characteristics. The varietycan be reproduced by planting and growing seeds of the variety underself-pollinating or sib-pollinating conditions, as is known to those ofskill in the agricultural arts. Soybean variety BK000496 shows novariants other than what would normally be expected due to environmentor that would occur for almost any characteristic during the course ofrepeated sexual reproduction. The variety description information (Table2) provides a summary of soybean variety BK000496 plant characteristics.Those of skill in the art will recognize that these are typical valuesthat may vary due to environment and that other values that aresubstantially equivalent are within the scope of the invention. As usedherein, “a soybean plant having the physiological and morphologicalcharacteristics of soybean variety BK000496” is a plant having thecharacteristics set forth in Table 2 when grown in the sameenvironmental conditions.

TABLE 2 Physiological and Morphological Characteristics of CultivarBK000496 Characteristic Value Relative Maturity 0.4 Hilum Color (MatureSeed) Imperfect Yellow Seed Coat Color (Mature Seed) Yellow CotyledonColor (Mature Seed) Yellow Canopy Width (1 to 9) 5 Growth HabitIndeterminate Plant Height (inches) 33 Lodging (1 to 5) 1.5 Flower ColorPurple Leaflet Shape Ovate Pubescence Color Tawny Pod Wall Color BrownResistance/Tolerance to Herbicides Roundup Ready (40-3-2) No RoundupReady 2 (GM_A19788) No Sulfonylurea Tolerant Soybean (STS) No

Soybean variety BK000496 in one embodiment of the present inventioncarries one or more transgenes, for example, a glyphosate tolerancetransgene, an auxin herbicide tolerance gene, a glufosinate tolerancegene, a desaturase gene or other transgenes. In another embodiment ofthe invention, the soybean does not carry any herbicide resistancetraits. In yet another embodiment of the invention, the soybean does notcarry any transgenes but carries alleles for aphid resistance, cystnematode resistance and/or disease resistance or the like. In stillanother embodiment, the soybean carries both alleles and transgenesproviding desired traits.

In addition to the individual plant characteristics set forth in Table2, agronomic yield of soybean variety BK000496 was evaluated. Table 3compares the yield and maturity difference of soybean variety BK000496to those of other soybean varieties developed for a similarcrop-production region.

TABLE 3 Yield of Cultivar BK000496 Compared to Selected CultivarsMaturity Years Paired Reps. Yield Yield Difference (#) Cultivart-test^(a) (#) (bu/ac) (%)^(b) (days) 1 BK000496 NS 14 41.68 100.0 0CL416933 44.76 107.4 10  1 BK000496 *** 14 41.68 100.0 0 CL416981 48.34116.0 8 1 BK000496 ** 14 41.68 100.0 0 CR0624 47.50 114.0 3 1 BK000496NS 14 41.68 100.0 0 CS09R202N 46.74 112.1 8 1 BK000496 NS 14 41.68 100.00 OACDRAYTON 44.34 106.4 4 ^(a)Thresholds for paired t-tests are nosignificant difference (NS) and significant at P < .05 (*), P < .01(**), and P < .001 (***). ^(b)Percentage yield relative to thelower-yielding cultivar in each two-way comparison.

In addition to the individual plant characteristics and yield data setforth in Tables 2 and 3, the food grade quality of soybean cultivarBK000496 was evaluated. BK000496 had satisfactory soymilk and tofumaking quality (Table 4). BK000496 is a food grade cultivar withexcellent agronomic characteristics including a yellow hilum, and highyield potential for the tofu and soymilk markets.

TABLE 4 Soymilk and Tofu Yield and Quality of BK000496 Grown in 2013Characteristic Value Water Uptake^(a) 2.55 Soymilk yield (L/kg protein)16.09 Soymilk yield (L/kg bean) 6.59 Solids content of soymilk (%) 10.85Tofu (GDL)^(b) yield (kg/kg protein) 16.09 Tofu (GDL)^(b) yield (kg/kgbean) 6.54 Tofu (GDL)^(b) compression hardness (N)^(c) 1.2174 Tofu(GDL)^(b) compression firmness (N/mm)^(d) 0.2368 Tofu (GDL)^(b)penetration hardness (N)^(c) 0.3141 Tofu (GDL)^(b) penetration firmness(N/mm)^(d) 0.0747 Tofu (CaSO₄)^(e) yield (kg/kg beans) 15.98 Tofu(CaSO₄)^(e) yield (kg/kg protein) 6.54 Tofu (CaSO₄)^(e) compressionhardness (N)^(c) 0.6877 Tofu (CaSO₄)^(e) compression firmness (N/mm)^(d)0.1441 Tofu (CaSO₄)^(e) penetration hardness (N)^(c) 0.2107 Tofu(CaSO₄)^(e) penetration firmness (N/mm)^(d) 0.0536 ^(a)(weight ofhydrated beans after soaking)/(initial weight of beans) ^(b)Tofu madewith Glucono-delta-lactone coagulant ^(c)Hardness in Newtons^(d)Firmness in Newtons per mm. ^(e)Tofu made with Calcium sulphatedihydrate coagulantSoybean Variety BK000496 Breeding and Production Methods

The present invention provides methods for producing soybean seed, orplants grown therefrom, by crossing the soybean variety BK000496 withitself or a second variety. These methods can be used for propagation ofthe soybean variety BK000496, or can be used to produce BK000496-derivedhybrid soybean seeds and the plants grown therefrom. Hybrid soybeanplants can be used in the commercial production of soy products or maybe advanced in certain breeding protocols for the production ofadditional novel soybean varieties by crossing the soybean varietyBK000496-derived soybean plant with itself or another soybean plant toyield an additional soybean variety BK000496-derived soybean progenyseed. This crossing process can be repeated one or more times togenerate additional soybean varieties. A hybrid plant can also be usedas a recurrent parent at any given stage in a backcrossing protocolduring the production of the soybean variety BK000496 which comprises anadded desired trait.

In some embodiments, the present invention provides for using theBK000496 soybean plant, or part thereof, or a soybean plant having thephysiological and morphological characteristics of the BK000496 soybeanplant, as a source of breeding material for developing anBK000496-derived soybean plant in a soybean breeding program using plantbreeding techniques. Plant breeding techniques useful in the developingsoybean plants include, but are not limited to, single seed descent,modified single seed descent, recurrent selection, reselection, massselection, bulk selection, backcrossing, pedigree breeding, mutationbreeding, restriction fragment length polymorphism enhanced selection,genetic marker enhanced selection, making double haploids andtransformation. Plant breeding techniques are known to the art and havebeen described in the literature. For example, see U.S. Pat. Nos.6,143,954; 7,803,996; and 7,807,884, which, along with the referencescited therein, is incorporated by reference herein.

Selection of soybean plants for breeding is not necessarily dependent onthe phenotype of a plant and instead can be based on geneticinvestigations. For example, one may utilize a suitable genetic markerwhich is closely genetically linked to a trait of interest. One of thesemarkers may therefore be used to identify the presence or absence of atrait in the offspring of a particular cross, and hence may be used inselection of progeny for continued breeding. This technique may commonlybe referred to as “marker assisted selection.” Any other type of geneticmarker or other assay which is able to identify the relative presence orabsence of a trait of interest in a plant may also be useful forbreeding purposes. Procedures for marker assisted selection applicableto the breeding of soybeans are well known in the art. Such methods willbe of particular utility in the case of recessive traits and variablephenotypes, or where conventional assays may be more expensive, timeconsuming or otherwise disadvantageous. Types of genetic markers whichcould be used in accordance with the invention include, but are notnecessarily limited to, Simple Sequence Length Polymorphisms (SSLPs),Restriction Fragment Length Polymorphisms (RFLPs), Randomly AmplifiedPolymorphic DNAs (RAPDs), DNA Amplification Fingerprinting (DAF),Sequence Characterized Amplified Regions (SCARs), Arbitrary PrimedPolymerase Chain Reaction (AP-PCR), Amplified Fragment LengthPolymorphisms (AFLPs) (EP 534 858, specifically incorporated herein byreference in its entirety), Simple Sequence Repeats (SSRs, also referredto as “Microsatellites”), and Single Nucleotide Polymorphisms (SNPs).

Many qualitative characters also have potential use as phenotype-basedgenetic markers in soybeans; however, some or many may not differ amongvarieties commonly used as parents. The most widely used genetic markersare flower color (purple dominant to white), pubescence color (browndominant to gray), and pod color (brown dominant to tan). Theassociation of purple hypocotyl color with purple flowers and greenhypocotyl color with white flowers is commonly used to identify hybridsin the seedling stage. Differences in maturity, height, hilum color,pubescence color, pod wall color, and pest resistance between parentscan also be used to verify hybrid plants.

Soybean variety BK000496 represents a novel base genetic variety intowhich a new desired locus or trait may be introduced by introgression.Backcrossing and direct transformation represent two important methodsthat can be used to accomplish such an introgression. In certainembodiments of the present invention, plants of soybean variety BK000496are provided modified to include one or more desired heritable traits.

Plants of the subject invention including one or more desired heritabletraits may, in one embodiment, be developed by a plant breedingtechnique called backcrossing, wherein essentially all of the desiredphysiological and morphological characteristics of a variety arerecovered, when grown in the same environmental conditions, in additionto a genetic locus comprising the desired trait transferred into theplant via the backcrossing technique. Backcrossing methods can be usedwith the present invention to improve or introduce a desired trait intosoybean variety BK000496. The parental soybean plant which contributesthe locus for the desired characteristic is termed the nonrecurrent ordonor parent. This terminology refers to the fact that the nonrecurrentparent is used one time in the backcross protocol and therefore does notrecur. The parental soybean plant (e.g., soybean variety BK000496) towhich the locus or loci from the nonrecurrent parent are transferred isknown as the recurrent parent as it is used for several rounds in thebackcrossing protocol (Poehlman et al., 1995; Fehr, 1987a,b; Sprague andDudley, 1988).

In a typical backcross protocol, the original variety of interest(recurrent parent, e.g., soybean variety BK000496) is crossed to asecond variety (nonrecurrent parent) that carries the single locus ofinterest to be transferred to produce F1 progeny plants. The resultingF1 progeny from this cross are then selected that have the desired traitand crossed again to the recurrent parent to produce backcross progenyplants having the desired trait and physiological and morphologicalcharacteristics of the recurrent parent. The process is repeated until asoybean plant is obtained wherein essentially all of the desiredmorphological and physiological characteristics of the recurrent parentare recovered in the converted plant, in addition to the transferredlocus comprising the desired trait from the nonrecurrent parent.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute a single or a very limited number of traits orcharacteristics into the original variety. To accomplish this, a singlelocus of the recurrent variety is modified or substituted with thedesired locus from the nonrecurrent parent, while retaining essentiallyall of the rest of the desired genetic, and therefore the desiredphysiological and morphological constitution of the original variety.The choice of the particular nonrecurrent parent will depend on thepurpose of the backcross; one of the major purposes is to add somecommercially desirable, agronomically important trait to the plant. Theexact backcrossing protocol will depend on the characteristic or traitbeing altered to determine an appropriate testing protocol. Althoughbackcrossing methods are simplified when the characteristic beingtransferred is a dominant allele, a recessive allele may also betransferred. In this instance it may be necessary to introduce a test ofthe progeny to determine if the desired characteristic has beensuccessfully transferred.

Soybean varieties can also be developed from more than two parents(Fehr, In: Soybeans: Improvement, Production and Uses, 2d Ed., Manograph16:249, 1987). The technique, known as modified backcrossing, usesdifferent recurrent parents during the backcrossing. Modifiedbackcrossing can be used to replace the original recurrent parent with avariety having certain more desirable characteristics or multipleparents may be used to obtain different desirable characteristics fromeach.

Many single locus traits have been identified that are not regularlyselected for in the development of a new inbred but that can be improvedby backcrossing techniques. Single locus traits may or may not betransgenic; examples of these traits include, but are not limited to,male sterility, herbicide resistance, insect resistance, modified fattyacid metabolism, modified carbohydrate metabolism and resistance tobacterial disease, fungal disease or viral disease.

Direct selection may be applied where the single locus acts as adominant trait. An example of a dominant trait is an herbicide tolerancetrait. For this selection process, the progeny of the initial cross aresprayed with the herbicide prior to the backcrossing. The sprayingeliminates any plants which do not have the desired herbicide tolerancecharacteristic, and only those plants which have the herbicide tolerancegene are used in the subsequent backcross. This process is then repeatedfor all additional backcross generations.

In other embodiments of the present invention, plants of the subjectinvention including one or more desired heritable traits may bedeveloped by direct transformation of soybean variety BK000496, orthrough the use of backcrossing approaches as described herein, forexample, to introgress a transgenic trait into soybean variety BK000496.Accordingly, in one embodiment of the present invention a method ofproducing a plant of soybean variety BK000496 comprising an addeddesired trait is provided, where the method comprises introducing atleast one transgene conferring the desired trait into variety BK000496.In some embodiments, introducing at least one transgene conferring thedesired trait comprises transforming a soybean plant, or part thereof,of variety BK000496 with one or more transgenes that confer at least onedesired trait. In another embodiment, introducing at least one transgeneconferring the desired trait comprises use of backcrossing to introgressa transgenic trait into soybean variety BK000496. Another embodiment isthe product produced by this process, wherein the product comprises theat least one desired trait and all of the physiological andmorphological characteristics of soybean variety BK000496 when grown inthe same location and in the same environmental conditions.

In one embodiment the desired trait may be one or more of malesterility, site-specific recombination, abiotic stress tolerance,herbicide tolerance, insect or pest resistance, disease resistance,fungal resistance, modified fatty acid metabolism, and modifiedcarbohydrate metabolism. The specific gene may be any known in the artor listed herein, including; a polynucleotide conferring resistance toglyphosate, phenoxyacetate auxins, pyridyloxyacetate auxins,phenoxyproprionate auxins, pehnoxybutanoate auxins, sulfonylurea,imidazalinone, dicamba, glufosinate, cyclohexone, triazine, andbenzonitrile; a polynucleotide encoding a Bacillus thuringiensispolypeptide, a polynucleotide encoding phytase, FAD-2, FAD-3, galactinolsynthase or a raffinose synthetic enzyme; or a polynucleotide conferringresistance to soybean cyst nematode, brown stem rot, phytophthora rootrot, soybean mosaic virus or sudden death syndrome.

Various genetic elements can be introduced into the plant genome usingtransformation. These elements include, but are not limited to genes;coding sequences; inducible, constitutive, and tissue specificpromoters; enhancing sequences; and signal and targeting sequences. Forexample, see the traits, genes and transformation methods listed in U.S.Pat. No. 6,118,055. The most prevalent types of plant transformationinvolve the construction of an expression vector. Such a vectorcomprises a DNA sequence that contains a gene under the control of oroperatively linked to a regulatory element, for example a promoter. Thevector may contain one or more genes and one or more regulatoryelements.

Plant transformation techniques which result in the generation oftransgenic plants are known in the art. These techniques include, butare not limited to:

(1) Projectile bombardment or microprojectile-mediated delivery. Thisprocedure involves propelling inert or biologically active particlescomplexed with DNA at plant cells, wherein the particles penetrate theouter surface of the cell and the DNA is incorporated within the genomeof the plant cell. See e.g., Klein et al., (1987) Nature 327: 70-73;Tomes et al., Plant Cell, Tissue & Organ Culture: Fundamental Methods,eds. Gambourg and Phillips (1995) (Springer-Velag, Berlin); Gordon-Kimet al., (1990) Plant Cell 2:603-618; U.S. Pat. Nos. 4,945,050,5,879,918, 5,932,782; 5,015,580, 5,550,318, 5,538,880, 6,160,208,6,399,861, and 6,403,865;

(2) Microinjection of plant cell protoplasts or embryogenic callus,including the use of silicon carbide mediated DNA uptake. See e.g.,Crossway et al., (1985) Molecular General Genetics 202:179; Kaeppler etal. (1990) Plant Cell Reporter 9:415-418;

(3) Direct gene transfer. See e.g., International Patent Application No.WO85/01856 and European Patent Application No. 0 275 069;

(4) Electroporation, calcium mediated, or PEG precipitation ofprotoplasts or cells comprising partial cell walls. See e.g., Fromm etal., (1985) Proceedings of the National Academy of Sciences 82: 5824;Paszkowski et al., (1984) European Molecular Biological Organization 3:2717-2722; Potrykus et al. (1985) Molecular General Genetics199:169-177; Shimamoto (1989) Nature 338:274-276; D'Halluin et al.(1992) Plant Cell 4: 1 495-1505.; U.S. Pat. No. 5,384,253;

(5) Aerosol beam technology employs the jet expansion of an inert gas asit passes from a region of higher gas pressure to a region of lower gaspressure through a small orifice. The expanding gas accelerates aerosoldroplets containing DNA molecules at supersonic speeds into a cell ortissue. See e.g., Held et al., U.S. Pat. Nos. 6,809,232; 7,067,716; and7,026,286.

(6) Agrobacterium-mediated transformations of plants are also included.Agrobacterium-mediated transformation is described in, for example,Horsch et al., (1984) Science 233:496-498, and Fraley et al., (1983)Proc. Nat. Acad. Sci. USA 80:4803 and U.S. Pat. Nos. 5,824,877;5,981,840, and 6,384,301; Ishida et al., (1996) Nature Biotechnology14:745-750. Generally, the Agrobacterium transformation system is usedto engineer dicotyledonous plants see Bevan et al (1982) Ann. Rev. Genet16:357-384; Rogers et al., (1986) Methods Enzymol. 118:627-641). TheAgrobacterium transformation system may also be used to transform, aswell as transfer, DNA to monocotyledonous plants and plant cells. SeeU.S. Pat. No. 5,591,616; Hemalsteen et al., (1984) EMBO J3:3039-3041;Hooykass-Van Slogteren et al., (1984) Nature 3 11: 763-764; Grimsley etal., (1987) Nature 325: 1677-179; Boulton et al., (1989) Plant Mol.Biol. 12:3 1-40.; and Gould et al., (1991) Plant Physiol. 95:426-434. Inaddition, gene transfer may be achieved using non-Agrobacterium bacteriaor viruses such as Rhizobium sp. NGR234, Sinorhizobium meliloti,Mesorhizobium loti, potato virus X, cauliflower mosaic virus and cassavavein mosaic virus and/or tobacco mosaic virus, See, e.g., Chung et al.,(2006) Trends Plant Sci. 11(1): 1-4; U.S. Pat. Nos. 6,660,500,6,462,255, 5,889,190 and 5,889,101.

Soybean transformation is described in particular in a number ofpublications. An example of an exemplary soybean transformationtechnique includes the use of Agrobacterium-mediated planttransformation. One example of soybean transformation comprisesinfecting half-seed explants of soybean with Agrobacterium tumefacienscontaining a transgene and regenerating the half-seed explants in vitroon selection medium. See U.S. Pat. No. 7,473,822 and Paz et al., (2006)Plant Cell Reports 25: 206-213. A second example ofAgrobacterium-mediated soybean transformation employs the use ofglufosinate as the selection system, thereby resulting in an enhancedtransformation efficiency. See Zeng et al., (2004) Plant Cell Rep22:478-482.

After effecting delivery of exogenous DNA to recipient cells, the nextsteps generally concern identifying the transformed cells for furtherculturing and plant regeneration. In order to improve the ability toidentify transformants, one may desire to employ a selectable orscreenable marker gene with the transformation vector used to generatethe transformant. In this case, the potentially transformed cellpopulation can be assayed by exposing the cells to a selective agent oragents, or the cells can be screened for the desired marker gene trait.

Cells that survive the exposure to the selective agent, or cells thathave been scored positive in a screening assay, may be cultured in mediathat supports regeneration of plants. In some embodiments, any suitableplant tissue culture media (e.g., MS and N6 media) may be modified byincluding further substances, such as growth regulators. Tissue may bemaintained on a basic media with growth regulators until sufficienttissue is available to begin plant regeneration efforts, or followingrepeated rounds of manual selection, until the morphology of the tissueis suitable for regeneration (e.g., at least 2 weeks), then transferredto media conducive to shoot formation. Cultures are transferredperiodically until sufficient shoot formation has occurred. Once shootsare formed, they are transferred to media conducive to root formation.Once sufficient roots are formed, plants can be transferred to soil forfurther growth and maturity.

To confirm the presence of a transgene in the regenerating plants, avariety of assays may be performed. Such assays include, for example:molecular biological assays, such as Southern and Northern blotting andPCR; biochemical assays, such as detecting the presence of a proteinproduct, e.g., by immunological means (e.g., ELISA and/or Western blots)or by enzymatic function; plant part assays, such as leaf or rootassays; and analysis of the phenotype of the whole regenerated plant.

Through the transformation of soybean, the expression of genes can bealtered to enhance disease resistance, insect resistance, herbicideresistance, agronomic, grain quality and other desired traits.Transformation can also be used to insert DNA sequences which control orhelp control male-sterility. DNA sequences native to soybean as well asnon-native DNA sequences can be transformed into soybean and used toalter levels of native or non-native proteins. Various promoters,targeting sequences, enhancing sequences, and other DNA sequences can beinserted into the genome for the purpose of altering the expression ofproteins. Reduction of the activity of specific genes (also known asgene silencing, or gene suppression) is desirable for several aspects ofgenetic engineering in plants.

Many techniques for gene silencing are well known to one of skill in theart, including but not limited to knock-outs (such as by insertion of atransposable element such as mu (Vicki Chandler, The Maize Handbook ch.118 (Springer-Verlag 1994) or other genetic elements such as a FRT, Loxor other site specific integration site, antisense technology (see,e.g., Sheehy et al. (1988) PNAS USA 85:8805-8809; and U.S. Pat. Nos.5,107,065; 5,453,566; and 5,759,829); co-suppression (e.g., Taylor(1997) Plant Cell 9:1245; Jorgensen (1990) Trends Biotech.8(12):340-344; Flavell (1994) PNAS USA 91:3490-3496; Finnegan et al.(1994) Bio/Technology 12: 883-888; and Neuhuber et al. (1994) Mol. Gen.Genet. 244:230-241); RNA interference (Napoli et al. (1990) Plant Cell2:279-289; U.S. Pat. No. 5,034,323; Sharp (1999) Genes Dev. 13:139-141;Zamore et al. (2000) Cell 101:25-33; and Montgomery et al. (1998) PNASUSA 95:15502-15507), virus-induced gene silencing (Burton, et al. (2000)Plant Cell 12:691-705; and Baulcombe (1999) Curr. Op. Plant Bio.2:109-113); target-RNA-specific ribozymes (Haseloff et al. (1988) Nature334: 585-591); hairpin structures (Smith et al. (2000) Nature407:319-320; WO 99/53050; and WO 98/53083); MicroRNA (Aukerman & Sakai(2003) Plant Cell 15:2730-2741); ribozymes (Steinecke et al. (1992) EMBOJ. 11:1525; and Perriman et al. (1993) Antisense Res. Dev. 3:253);oligonucleotide-mediated targeted modification (e.g., WO 03/076574 andWO 99/25853); Zn-finger targeted molecules (e.g., WO 01/52620; WO03/048345; and WO 00/42219); and other methods or combinations of theabove methods known to those of skill in the art.

Exemplary nucleotide sequences or encoded polypeptides that may bealtered or introduced by genetic engineering to provide desired traitsinclude, but are not limited to, those categorized below.

1. Genes or Encoded Proteins that Confer Resistance to Pests or Disease

(A) Plant Disease Resistance Genes. Plant defenses are often activatedby specific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant variety can be transformed with clonedresistance gene to engineer plants that are resistant to specificpathogen strains. Examples of such genes include, the tomato Cf-9 genefor resistance to Cladosporium flavum (Jones et al., 1994 Science266:789), tomato Pto gene, which encodes a protein kinase, forresistance to Pseudomonas syringae pv. tomato (Martin et al., 1993Science 262:1432), and Arabidopsis RSSP2 gene for resistance toPseudomonas syringae (Mindrinos et al., 1994 Cell 78:1089).

(B) A Bacillus thuringiensis protein, a derivative thereof or asynthetic polypeptide modeled thereon, such as, a nucleotide sequence ofa Bt δ-endotoxin gene (Geiser et al., 1986 Gene 48:109). Moreover, DNAmolecules encoding δ-endotoxin genes can be purchased from American TypeCulture Collection (Rockville, Md.), under ATCC accession numbers.40098, 67136, 31995 and 31998. Other non-limiting examples of Bacillusthuringiensis transgenes being genetically engineered are given in thefollowing patents, patent applications and publications and hereby areincorporated by reference for this purpose: U.S. Pat. Nos. 5,188,960;5,689,052; 5,880,275; 5,986,177; 7,105,332; 7,208,474; WO 91/14778; WO99/31248; WO 01/12731; WO 99/24581; WO 97/40162 and U.S. applicationSer. Nos. 10/032,717; 10/414,637; 11/018,615; 11/404,297; 11/404,638;11/471,878; 11/780,501; 11/780,511; 11/780,503; 11/953,648; 11/953,648;and 11/957,893, and Estruch, et al., 1996. Proc. Natl. Acad. Sci.93:5389.

(C) A lectin, such as, nucleotide sequences of several Clivia miniatamannose-binding lectin genes (Van Damme et al., 1994 Plant Molec. Biol.24:825).

(D) A vitamin binding protein, such as avidin and avidin homologs whichare useful as larvicides against insect pests. See U.S. Pat. No.5,659,026.

(E) An enzyme inhibitor, e.g., a protease inhibitor or an amylaseinhibitor. Examples of such genes include, a rice cysteine proteinaseinhibitor (Abe et al., 1987 J. Biol. Chem. 262:16793), a tobaccoproteinase inhibitor I (Huub et al., 1993 Plant Molec. Biol. 21:985),and an α-amylase inhibitor Sumitani et al., 1993 Biosci. Biotech.Biochem. 57:1243).

(F) An insect-specific hormone or pheromone such as an ecdysteroid andjuvenile hormone a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof, such as, baculovirus expression of clonedjuvenile hormone esterase, an inactivator of juvenile hormone (Hammocket al., 1990 Nature 344:458).

(G) An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. Examples of such genesinclude, an insect diuretic hormone receptor (Regan, 1994), anallostatin identified in Diploptera punctata (Pratt, 1989),insect-specific, paralytic neurotoxins (U.S. Pat. No. 5,266,361).

(H) An insect-specific venom produced in nature by a snake, a wasp,etc., such as, a scorpion insectotoxic peptide (Pang, 1992 Gene116:165).

(I) An enzyme responsible for a hyperaccumulation of monoterpene, asesquiterpene, a steroid, hydroxamic acid, a phenylpropanoid derivativeor another non-protein molecule with insecticidal activity.

(J) An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme, anuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase and a glucanase, whether natural or synthetic. Examples ofsuch genes include, a callas gene (PCT published applicationWO93/02197), chitinase-encoding sequences (which can be obtained, forexample, from the ATCC under accession numbers 3999637 and 67152),tobacco hookworm chitinase (Kramer et al., 1993 Insect Molec. Biol.23:691) and parsley ubi4-2 polyubiquitin gene (Kawalleck et al., 1993Plant Molec. Biol. 21:673).

(K) A molecule that stimulates signal transduction. Examples of suchmolecules include, nucleotide sequences for mung bean calmodulin cDNAclones (Botella et al., 1994 Plant Molec. Biol. 24:757) and a nucleotidesequence of a maize calmodulin cDNA clone (Griess et al., 1994 PlantPhysiol. 104:1467).

(L) A hydrophobic moment peptide. See U.S. Pat. Nos. 5,659,026 and5,607,914, the latter teaches synthetic antimicrobial peptides thatconfer disease resistance.

(M) A membrane permease, a channel former or a channel blocker, such as,a cecropin-β lytic peptide analog (Jaynes et al., 1993 Plant Sci. 89:43)which renders transgenic tobacco plants resistant to Pseudomonassolanacearum.

(N) A viral-invasive protein or a complex toxin derived there from. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. Coat protein-mediated resistance has beenconferred upon transformed plants against alfalfa mosaic virus, cucumbermosaic virus, tobacco streak virus, potato virus X, potato virus Y,tobacco etch virus, tobacco rattle virus and tobacco mosaic virus. See,for example, Beachy et al. (1990) Ann. Rev. Phytopathol. 28:451.

(O) An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect. Forexample, Taylor et al. (1994) Abstract #497, Seventh Int'l. Symposium onMolecular Plant-Microbe Interactions, shows enzymatic inactivation intransgenic tobacco via production of single-chain antibody fragments.

(P) A virus-specific antibody. See, for example, Tavladoraki et al.(1993) Nature 266:469, which shows that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

(Q) A developmental-arrestive protein produced in nature by a pathogenor a parasite. Thus, fungal endo α-1,4-D polygalacturonases facilitatefungal colonization and plant nutrient release by solubilizing plantcell wall homo-α-1,4-D-galacturonase (Lamb et al., 1992) Bio/Technology10:1436. The cloning and characterization of a gene which encodes a beanendopolygalacturonase-inhibiting protein is described by Toubart et al.(1992 Plant J. 2:367).

(R) A developmental-arrestive protein produced in nature by a plant,such as, the barley ribosome-inactivating gene has an increasedresistance to fungal disease (Longemann et al., 1992). Bio/Technology10:3305.

(S) A small RNA (e.g., antisense, hairpin, siRNA, or miRNA) thatinhibits expression of a pathogen gene necessary for the pathogen tosurvive or thrive.

2. Genes that Confer Resistance to a Herbicide

(A) Genes encoding resistance or tolerance to a herbicide that inhibitsthe growing point or meristem, such as an imidazalinone or asulfonylurea. Exemplary genes in this category code for mutant ALS (Leeet al., 1988 EMBOJ. 7:1241) and AHAS enzyme (Miki et al., 1990 Theor.Appl. Genet. 80:449).

(B) One or more additional genes encoding resistance or tolerance toglyphosate imparted by mutant EPSP synthase and aroA genes, or throughmetabolic inactivation by genes such as GAT (glyphosateacetyltransferases or GOX (glyphosate oxidase) and other phosphonocompounds such as glufosinate (PAT and bar genes), and pyridinoxy orphenoxy proprionic acids and cyclohexones (ACCase inhibitor encodinggenes). See, for example, U.S. Pat. No. 4,940,835, which discloses thenucleotide sequence of a form of EPSP which can confer glyphosateresistance. A DNA molecule encoding a mutant aroA gene can be obtainedunder ATCC accession number 39256, and the nucleotide sequence of themutant gene is disclosed in U.S. Pat. No. 4,769,061. European patentapplication No. 0 333 033 and U.S. Pat. No. 4,975,374 disclosenucleotide sequences of glutamine synthetase genes which conferresistance to herbicides such as L-phosphinothricin. The nucleotidesequence of a phosphinothricinacetyl-transferase gene is provided inEuropean application No. 0 242 246. De Greef et al. (1989)Bio/Technology 7:61 describes the production of transgenic plants thatexpress chimeric bar genes coding for phosphinothricin acetyltransferase activity. Exemplary genes conferring resistance to phenoxyproprionic acids and cyclohexones, such as sethoxydim and haloxyfop, arethe Accl-S1, Accl-S2 and Accl-S3 genes described by Marshall et al.(1992) Theor. Appl. Genet. 83:435.

(C) Genes encoding resistance or tolerance to a herbicide that inhibitsphotosynthesis, such as a triazine (psbA and gs+ genes) and abenzonitrile (nitrilase gene). Przibilla et al. (1991) Plant Cell 3:169describes the use of plasmids encoding mutant psbA genes to transformChlamydomonas. Nucleotide sequences for nitrilase genes are disclosed inU.S. Pat. No. 4,810,648, and DNA molecules containing these genes areavailable under ATCC accession numbers 53435, 67441 and 67442. Cloningand expression of DNA coding for a glutathione S-transferase isdescribed by Hayes et al. (1992) Biochem. J. 285:173.

(D) Genes encoding resistance or tolerance to a herbicide that bind tohydroxyphenylpyruvate dioxygenases (HPPD), enzymes which catalyze thereaction in which para-hydroxyphenylpyruvate (HPP) is transformed intohomogentisate. This includes herbicides such as isoxazoles (EP418175,EP470856, EP487352, EP527036, EP560482, EP682659, U.S. Pat. No.5,424,276), in particular isoxaflutole, which is a selective herbicidefor maize, diketonitriles (EP496630, EP496631), in particular2-cyano-3-cyclopropyl-1-(2-SO2CH3-4-CF3 phenyl)propane-1,3-dione and2-cyano-3-cyclopropyl-1-(2-SO2CH3-4-2,3Cl2phenyl)propane-1,3-dione,triketones (EP625505, EP625508, U.S. Pat. No. 5,506,195), in particularsulcotrione, or else pyrazolinates. A gene that produces anoverabundance of HPPD in plants can provide tolerance or resistance tosuch herbicides, including, for example, genes described at U.S. Pat.Nos. 6,268,549 and 6,245,968 and US publication No. 20030066102.

(E) Genes encoding resistance or tolerance to phenoxyacetate auxinherbicides, such as 2,4-dichlorophenoxyacetic acid (2,4-D) and which mayalso confer resistance or tolerance to “fop” herbicides. Examples ofsuch genes include the α-ketoglutarate-dependent dioxygenase enzyme(AAD-1) gene, described at US Patent Publication 20090093366.

(F) Genes encoding resistance or tolerance to phenoxyacetate auxinherbicides, such as 2,4-dichlorophenoxyacetic acid (2,4-D) and which mayalso confer resistance or tolerance to pyridyloxyacetate auxins (such asfluroxypyr and triclopyr), phenoxyproprionate auxins (such as MCPA anddichloprop), pehnoxybutanoate auxins (such as 2,4-DB). Examples of suchgenes include the α-ketoglutarate-dependent dioxygenase enzyme (AAD-12)gene, described in WO 2007/053482 A2.

(G) Genes encoding resistance or tolerance to dicamba, such as dicambamonooxygenase (DMO) from Pseudomonas maltophilia which is involved inthe conversion of a herbicidal form of the herbicide dicamba to anon-toxic 3,6-dichlorosalicylic acid and thus may be used for producingplants tolerant to this herbicide. See, e.g., US Patent Application No:20030135879.

3. Genes that Confer or Contribute to a Value-Added Trait

(A) Modified fatty acid metabolism, for example, by transforming maizeor Brassica with a small RNA or stearoyl-ACP desaturase to increasestearic acid content of the plant (Knultzon et al., 1992) Proc. Nat.Acad. Sci. USA 89:2624.

(B) Decreased phytate content

(1) Introduction of a phytase-encoding gene would enhance breakdown ofphytate, adding more free phosphate to the transformed plant, such asthe Aspergillus niger phytase gene (Van Hartingsveldt et al., 1993 Gene127:87).

(2) A gene could be introduced that reduces phytate content. In maize,this, for example, could be accomplished by cloning and thenreintroducing DNA associated with the single allele which is responsiblefor maize mutants characterized by low levels of phytic acid (Raboy etal., 1990 Maydica 35:383).

(C) Modified carbohydrate composition effected, for example, bytransforming plants with a gene coding for an enzyme that alters thebranching pattern of starch. Examples of such enzymes include,Streptococcus mucus fructosyltransferase gene (Shiroza et al., 1988) J.Bacteriol. 170:810, Bacillus subtilis levansucrase gene (Steinmetz etal., 1985 Mol. Gen. Genel. 200:220), Bacillus licheniformis α-amylase(Pen et al., 1992 Bio/Technology 10:292), tomato invertase genes (Elliotet al., 1993), barley amylase gene (Sogaard et al., 1993 J. Biol. Chem.268:22480), and maize endosperm starch branching enzyme II (Fisher etal., 1993 Plant Physiol. 102:1045).

(D) Altered antioxidant content or composition, such as alteration oftocopherol or tocotrienols. For example, see U.S. Pat. No. 6,787,683,US2004/0034886 and WO 00/68393 involving the manipulation of antioxidantlevels, and WO 03/082899 through alteration of a homogentisate geranylgeranyl transferase (hggt).

(E) Altered essential seed amino acids. For example, see U.S. Pat. No.6,127,600 (method of increasing accumulation of essential amino acids inseeds), U.S. Pat. No. 6,080,913 (binary methods of increasingaccumulation of essential amino acids in seeds), U.S. Pat. No. 5,990,389(high lysine), WO 99/40209 (alteration of amino acid compositions inseeds), WO 99/29882 (methods for altering amino acid content ofproteins), U.S. Pat. No. 5,850,016 (alteration of amino acidcompositions in seeds), WO 98/20133 (proteins with enhanced levels ofessential amino acids), U.S. Pat. No. 5,885,802 (high methionine), U.S.Pat. No. 5,885,801 (high threonine), U.S. Pat. No. 6,664,445 (plantamino acid biosynthetic enzymes), U.S. Pat. No. 6,459,019 (increasedlysine and threonine), U.S. Pat. No. 6,441,274 (plant tryptophansynthase beta subunit), U.S. Pat. No. 6,346,403 (methionine metabolicenzymes), U.S. Pat. No. 5,939,599 (high sulfur), U.S. Pat. No. 5,912,414(increased methionine), WO 98/56935 (plant amino acid biosyntheticenzymes), WO 98/45458 (engineered seed protein having higher percentageof essential amino acids), WO 98/42831 (increased lysine), U.S. Pat. No.5,633,436 (increasing sulfur amino acid content), U.S. Pat. No.5,559,223 (synthetic storage proteins with defined structure containingprogrammable levels of essential amino acids for improvement of thenutritional value of plants), WO 96/01905 (increased threonine), WO95/15392 (increased lysine), US2003/0163838, US2003/0150014,US2004/0068767, U.S. Pat. No. 6,803,498, WO 01/79516.

4. Genes that Control Male-Sterility

There are several methods of conferring genetic male sterilityavailable, such as multiple mutant genes at separate locations withinthe genome that confer male sterility, as disclosed in U.S. Pat. Nos.4,654,465 and 4,727,219 to Brar et al. and chromosomal translocations asdescribed by Patterson in U.S. Pat. Nos. 3,861,709 and 3,710,511. Inaddition to these methods, Albertsen et al., U.S. Pat. No. 5,432,068,describe a system of nuclear male sterility which includes: identifyinga gene which is critical to male fertility; silencing this native genewhich is critical to male fertility; removing the native promoter fromthe essential male fertility gene and replacing it with an induciblepromoter; inserting this genetically engineered gene back into theplant; and thus creating a plant that is male sterile because theinducible promoter is not “on” resulting in the male fertility gene notbeing transcribed. Fertility is restored by inducing, or turning “on”,the promoter, which in turn allows the gene that confers male fertilityto be transcribed.

(A) Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN—Ac-PPT (WO 01/29237).

(B) Introduction of various stamen-specific promoters (WO 92/13956, WO92/13957).

(C) Introduction of the barnase and the barstar gene (Paul et al. PlantMol. Biol. 19:611-622, 1992).

For additional examples of nuclear male and female sterility systems andgenes, see also, U.S. Pat. Nos. 5,859,341; 6,297,426; 5,478,369;5,824,524; 5,850,014; and 6,265,640; all of which are herebyincorporated by reference.

5. Genes that affect abiotic stress resistance (including but notlimited to enhancement of nitrogen utilization efficiency, alterednitrogen responsiveness, drought resistance or tolerance, coldresistance or tolerance, and salt resistance or tolerance) and increasedyield under stress. For example, see: WO 00/73475 where water useefficiency is altered through alteration of malate; U.S. Pat. Nos.5,892,009, 5,965,705, 5,929,305, 5,891,859, 6,417,428, 6,664,446,6,706,866, 6,717,034, 6,801,104, WO 00/060089, WO 01/026459, WO01/035725, WO 01/034726, WO 01/035727, WO 01/036444, WO 01/036597, WO01/036598, WO 02/015675, WO 02/017430, WO 0/2077185, WO 02/079403, WO03/013227, WO 03/013228, WO 03/014327, WO 04/031349, WO 04/076638, WO98/09521, and WO 99/38977 describing genes, including CBF genes andtranscription factors effective in mitigating the negative effects offreezing, high salinity, and drought on plants, as well as conferringother positive effects on plant phenotype; US2004/0148654 and WO01/36596 where abscisic acid is altered in plants resulting in improvedplant phenotype such as increased yield and/or increased tolerance toabiotic stress; WO 00/006341, WO 04/090143, U.S. application Ser. Nos.10/817,483 and 09/545,334 where cytokinin expression is modifiedresulting in plants with increased stress tolerance, such as droughttolerance, and/or increased yield. Also see WO 02/02776, WO 03/052063,JP2002281975, U.S. Pat. No. 6,084,153, WO 01/64898, U.S. Pat. No.6,177,275, and U.S. Pat. No. 6,107,547 (enhancement of nitrogenutilization and altered nitrogen responsiveness). For ethylenealteration, see US2004/0128719, US2003/0166197 and WO 00/32761. Forplant transcription factors or transcriptional regulators of abioticstress, see e.g. US2004/0098764 or US2004/0078852.

Other genes and transcription factors that affect plant growth andagronomic traits such as yield, flowering, plant growth and/or plantstructure, can be introduced or introgressed into plants, see e.g. WO97/49811 (LHY), WO 98/56918 (ESD4), WO 97/10339 and U.S. Pat. No.6,573,430 (TFL), U.S. Pat. No. 6,713,663 (FT), WO 96/14414 (CON), WO96/38560, WO 01/21822 (VRN1), WO 00/44918 (VRN2), WO 99/49064 (GI), WO00/46358 (FRI), WO 97/29123, U.S. Pat. No. 6,794,560, U.S. Pat. No.6,307,126 (GAI), WO 99/09174 (D8 and Rht), and WO 04/076638 and WO04/031349 (transcription factors).

One may obtain soybean plants according to the present invention bydirectly growing the seed of BK000496 or by any other means. A soybeanplant having all of the physiological and morphological characteristicsof BK000496 can be obtained by any suitable means, including, but notlimited to, regenerating plants or plant parts from tissue culture orcuttings. The scope of the present invention is not limited by themethod by which the plant is obtained.

Tissue Cultures and Plants Regenerated Therefrom

The present invention provides a tissue culture of protoplasts orregenerable cells from a plant, or parts thereof, produced from soybeanvariety BK000496, or a part thereof. In some embodiments, theprotoplasts or regenerable cells are derived from embryo, meristematiccell, leaf, pollen, ovule, cotyledon, hypocotyl, embryo, root, root tip,anther, pistil, pod, flower, shoot or stalk of soybean variety BK000496.

Tissue culture of various tissues of soybeans and regeneration of plantstherefrom is well known and widely published. For example, reference maybe had to Komatsuda, T. et al., “Genotype X Sucrose Interactions forSomatic Embryogenesis in Soybean,” Crop Sci. 31:333-337 (1991);Stephens, P. A. et al., “Agronomic Evaluation of Tissue-Culture-DerivedSoybean Plants,” Theor. Appl. Genet. (1991) 82:633-635; Komatsuda, T. etal., “Maturation and Germination of Somatic Embryos as Affected bySucrose and Plant Growth Regulators in Soybeans Glycine gracilis Skvortzand Glycine max (L.) Merr.,” Plant Cell, Tissue and Organ Culture,28:103-113 (1992); Dhir, S. et al., “Regeneration of Fertile Plants fromProtoplasts of Soybean (Glycine max L. Men.): Genotypic Differences inCulture Response,” Plant Cell Reports (1992) 11:285-289; Pandey, P. etal., “Plant Regeneration from Leaf and Hypocotyl Explants of Glycinewightii (W. and A.) VERDC. var. longicauda,” Japan J. Breed. 42:1-5(1992); and Shetty, K., et al., “Stimulation of In Vitro ShootOrganogenesis in Glycine max (Merrill.) by Allantoin and Amides,” PlantScience 81:(1992) 245-251; as well as U.S. Pat. No. 5,024,944, issuedJun. 18, 1991 to Collins et al. and U.S. Pat. No. 5,008,200, issued Apr.16, 1991 to Ranch et al., the disclosures of which are herebyincorporated herein in their entirety by reference. Thus, another aspectof the present invention is to provide cells which upon growth anddifferentiation produce soybean plants having the physiological andmorphological characteristics of soybean variety BK000496.

Soybean Products

Soybean is useful not only as a seed for producing soybean plants, butalso has utility as a grain. The grain can be used as a food source forboth animals and humans. Soybean is widely used as a source of proteinfor animal feeds for poultry, swine and cattle. The soybean grain istherefore a commodity. The soybean commodity plant products include butare not limited to protein concentrate, protein isolate, soybean hulls,meal, flower, oil and the whole soybean itself.

During processing of whole soybeans, the fibrous hull is removed and theoil is extracted. The remaining soybean meal is a combination ofcarbohydrates and approximately 50% protein. For human or animalconsumption soybean meal is made into soybean flour that is processed toprotein concentrates used for meat extenders or specialty pet foods.Production of edible protein ingredients from soybean offers a healthyless expensive replacement for animal protein in meats as well asdairy-type products. Soybeans and oils of BLT090009 can be suitable foruse in a variety of soyfoods made from whole soybeans, such as soymilk,soy nut butter, natto, and tempeh, and soyfoods made from processedsoybeans and soybean oil, including soybean meal, soy flour, soy proteinconcentrate, soy protein isolates, texturized soy protein concentrate,hydrolyzed soy protein, whipped topping, cooking oil, salad oil,shortening, and lecithin. Whole soybeans are also edible, and aretypically sold to consumers raw, roasted, or as edamame. Soymilk, whichis typically produced by soaking and grinding whole soybeans, may beconsumed without other processing, spray-dried, or processed to form soyyogurt, soy cheese, tofu, or yuba.

Accordingly, the present invention includes in some embodiments methodsfor producing a commodity plant product, which comprise obtaining seedof soybean variety BK000496 and producing the commodity plant productsdisclosed above. The invention further comprises soybean commodity plantproducts derived from soybean variety BK000496 seed according to thesemethods.

Deposit Information

Seed from soybean variety BK000496, disclosed above and recited in theappended claims, was deposited with the American Type Culture Collection(ATCC), 10801 University Boulevard, Manassas, Va. 20110 on Sep. 1, 2015under the Accession No. PTA-122489. The seeds deposited were taken fromseeds maintained by Dairyland Seed Co., Inc., West Bend, Wis. 53095since prior to the filing date of this application. Access to the ATCCdeposit will be available during the pendency of the application to theCommissioner of Patents and Trademarks and persons determined by theCommissioner to be entitled thereto upon request. Upon allowance of anyclaims in the application, the Applicant will make the depositirrevocably available to the public pursuant to 37 C.F.R. §1.808.Applicant has or will have satisfied all of the requirements of 37C.F.R. §§1.801-1.809. The deposit will be maintained in the ATCCdepository for a period of 30 years, or 5 years after the last request,or for the effective life of the patent, whichever is longer, and willbe replaced if necessary during that period.

All publications, patents and patent applications referenced in thespecification are indicative of the level of those skilled in the art towhich this invention pertains. All such publications, patents and patentapplications are incorporated by reference herein to the same extent asif each was specifically and individually indicated to be incorporatedby reference herein.

The foregoing invention has been described in some detail by way ofillustration and example for purposes of clarity and understanding.However, it should be appreciated by those having ordinary skill in theart that certain changes and modifications such as single genemodifications and mutations, somoclonal variants, variant individualsselected from large populations of the plants of the instant novelvariety and the like may be practiced within the scope of the invention,as limited only by the scope of the appended claims, without departingfrom the true concept, spirit, and scope of the invention.

What is claimed is:
 1. A seed of soybean variety BK000496, wherein arepresentative sample of the seed having been deposited under ATCCAccession No. PTA-122489.
 2. A plant of soybean variety BK000496, or apart thereof, wherein representative seed of said soybean varietyBK000496 having been deposited under ATCC Accession Number PTA-122489.3. The plant part of claim 2, wherein the part is a pollen grain, anovule, a protoplast, a cell, an embryo, a cotyledon, a hypocotyl, ameristem, a root, a pistil, an anther, a flower, a stem, a pod, a leafor a petiole.
 4. A soybean plant, or a part thereof, having all thephysiological and morphological characteristics of the soybean plant ofclaim 2 when grown in the same environmental conditions.
 5. A tissueculture of protoplasts or regenerable cells from the plant, of claim 2.6. The tissue culture of protoplast or regenerable cells of claim 5,wherein the protoplasts or regenerable cells are derived from embryo,meristematic cell, leaf, pollen, ovule, cotyledon, hypocotyl, root, roottip, anther, pistil, pod, flower, shoot or stalk.
 7. A soybean plantregenerated from the tissue culture of claim 6, wherein the plant hasall of the physiological and morphological characteristics of a plantproduced by growing seed of soybean variety BK000496, a representativesample of the seed having been deposited under ATCC Accession No.PTA-122489.
 8. A method for producing a soybean seed, comprisingcrossing soybean plants and harvesting the resultant seed, wherein atleast one soybean plant is the soybean plant of claim
 2. 9. An F1soybean seed produced by the method of claim
 8. 10. An F1 soybean plantproduced by growing the seed of claim
 9. 11. A method for producing asoybean variety BK000496-derived soybean plant, comprising: (a) crossinga soybean variety BK000496 plant of claim 2 with a second soybean plantto yield progeny soybean seed; and (b) growing said progeny seed toyield a soybean variety BK000496-derived soybean plant.
 12. An F1soybean variety BK000496-derived soybean plant produced by the method ofclaim
 11. 13. The method of claim 11, further comprising (c) crossingthe soybean variety BK000496-derived soybean plant of (b) with itself oranother soybean plant to yield an additional soybean varietyBK000496-derived soybean progeny seed; and (d) growing the progenysoybean seed of (c) to yield additional soybean variety BK000496-derivedsoybean plants.
 14. The method of claim 13, wherein (c) and (d) arerepeated at least one time to generate additional soybean varietyBK000496-derived soybean plants.
 15. A method of producing a plant ofsoybean variety BK000496 comprising an added desired trait, the methodcomprising introducing at least one transgene or locus conferring thedesired trait into the plant of claim
 2. 16. The method of claim 15,wherein the desired trait is selected from the group consisting of malesterility, site-specific recombination, abiotic stress tolerance,herbicide tolerance, insect or pest resistance, disease resistance,fungal resistance, modified fatty acid metabolism, and modifiedcarbohydrate metabolism.
 17. The method of claim 16, wherein the desiredtrait is herbicide tolerance and the tolerance is conferred to one ormore herbicides selected from the group consisting of glyphosate,phenoxyacetate auxins, pyridyloxyacetate auxins, phenoxyproprionateauxins, pehnoxybutanoate auxins, sulfonylurea, imidazalinone, dicamba,glufosinate, cyclohexone, triazine, and benzonitrile.
 18. The method ofclaim 16, wherein the desired trait is insect resistance and thetransgene encodes a Bacillus thuringiensis (Bt) endotoxin.
 19. A plantproduced by the method of claim 15, wherein the plant comprises thedesired trait and all of the physiological and morphologicalcharacteristics of soybean variety BK000496 when grown in the samelocation and in the same environmental conditions, and wherein arepresentative sample of seed of said soybean variety BK000496 havingbeen deposited under ATCC Accession No. PTA-122489.
 20. A method ofproducing a progeny soybean variety derived from variety BK000496comprising a desired trait, comprising: (a) crossing a soybean varietyBK000496 plant of claim 2 with a plant of another soybean variety thatcomprises a desired trait to produce an F1 progeny plant; (b) selectingone or more F1 progeny plants that have the desired trait to produceselected progeny plants; (c) crossing the selected progeny plants withthe soybean variety BK000496 plant to produce backcross progeny plants;(d) selecting for backcross progeny plants that have the desired traitand physiological and morphological characteristics of the soybeanvariety BK000496 plant to produce selected backcross progeny plants; and(e) repeating steps (c) and (d) a sufficient number of times insuccession to produce selected second or higher backcross progeny plantsthat comprise the desired trait and the physiological and morphologicalcharacteristics of soybean variety BK000496 when grown in the sameenvironmental conditions.
 21. The method of claim 20, wherein thedesired trait is selected from the group consisting of male sterility,herbicide resistance, insect resistance, modified fatty acid metabolism,modified carbohydrate metabolism and resistance to bacterial disease,fungal disease or viral disease.
 22. A plant produced by the method ofclaim 20, wherein the plant has the desired trait and all of the otherphysiological and morphological characteristics of soybean varietyBK000496 when grown in the same environmental conditions, and wherein arepresentative sample of seed of said soybean variety BK000496 havingbeen deposited under ATCC Accession No. PTA-122489.
 23. A method ofproducing a commodity plant product comprising obtaining the plant ofclaim 2, or a part thereof, and producing said commodity plant producttherefrom.
 24. The method of claim 23, wherein the commodity plantproduct is soybean meal, soy flour, soy protein concentrate, soy proteinisolates, texturized soy protein concentrate, hydrolyzed soy protein,whipped topping, whole soy seed, processed soy seed, animal feed,soymilk, soy nut butter, natto, tempeh, edible raw whole soybean pods,roasted whole soybean pods, edamame, soymilk, soy yogurt, soy cheese,tofu, yuba, and biomass.